1. Field of Invention
The present invention relates generally to assays, and more specifically to a lateral flow assay device with improved detection access.
2. Discussion of Related Art
Various chromatographic immunoassay techniques have been available for many years. Tests that can be performed with such chromatographic systems are, among others, immunoassays, which depend on the specific interaction between an antigen or hapten and a corresponding antibody. Immunoassays have been used as a means of testing for the presence or amount, or both, of clinically important molecules for some time. Immune-based latex agglutination tests for detecting a factor associated with rheumatoid arthritis were used as early as 1956 (Singer et al., Am. J. Med. 22:888-892 (1956)).
Among the many analytical systems used for detection of analytes, particularly analytes of biological interest, are chromatographic assay systems. Among the analytes frequently assayed with such systems are: (1) hormones, such as human chorionic gonadotropin (hCG), which is frequently assayed as a marker of human pregnancy; (2) antigens, particularly antigens specific to bacterial, viral, and protozoan pathogens, such as streptococcus, hepatitis virus, and giardia; (3) antibodies, particularly antibodies induced as a result of infection with pathogens, such as antibodies to the bacterium HELICOBACTER pylori and to Human Immunodeficiency Virus (HIV); (4) other proteins, such as hemoglobin, frequently assayed in determinations of fecal occult blood, an early indicator of gastrointestinal disorders such as colon cancer; (5) enzymes, such as aspartate aminotransferase, lactate dehydrogenase, alkaline phosphatase, and glutamate dehydrogenase, frequently assayed as indicators of physiological function and tissue damage; (6) drugs, both therapeutic drugs, such as antibiotics, tranquilizers and anticonvulsants, and illegal drugs of abuse, such as cocaine, heroin and marijuana; (7) vitamins; and (8) nucleic acid material.
Such chromatographic systems are frequently used by physicians and medical technicians for rapid in-office diagnosis, commonly referred to as “point of care” (POC) devices, and therapeutic monitoring of a variety of conditions and disorders. They are also increasingly used by: patients themselves for at-home monitoring of such conditions and disorders; scientists for use in field testing for transgenic crops and environmental contaminates; soldiers in battlefield conditions for biological warfare weapon detection; and veterinary and emergency technicians for rapid testing, among others.
Included in the chromatographic techniques used in conjunction with immunoassays is a procedure known as immunochromatography. In general, this technique uses a labeling reagent or particle that has been linked to an antibody for the molecule to be assayed, forming a conjugate. This conjugate is then mixed with a specimen and, if the molecule to be assayed is present in the specimen, the labeling reagent-linked antibodies bind to the molecule to be assayed, thereby giving an indication that the molecule to be assayed is present. The labeling reagent or particle can be identifiable by color, magnetic properties, radioactivity, specific reactivity with another molecule, or another physical or chemical property. The specific reactions that are employed vary with the nature of the molecule being assayed and the sample to be tested.
Immunochromatographic assays fall into two principal categories: “sandwich” and “competitive,” according to the nature of the antigen-antibody complex to be detected and the sequence of reactions required to produce that complex. In the case of antigen detection, the sandwich immunochromatographic procedures call for mixing the sample that may contain the analyte to be assayed with antibodies to the analyte. These antibodies are mobile and typically are linked to a label or a reagent, such as dyed latex, a colloidal metal sol, or a radioisotope. This mixture is then applied to a chromatographic medium containing a band or capture zone. This band or capture zone contains immobilized antibodies for the analyte of interest. The chromatographic medium can also be in the form of a strip resembling a dipstick. When the complex of the molecule to be assayed and the labeled antibody reaches the zone of the immobilized antibodies on the chromatographic medium, binding occurs, and the bound-labeled antibodies are localized at the zone. This indicates the presence of the molecule to be assayed. This technique can be used to obtain qualitative results. Examples of sandwich immunoassays performed on test strips are described in U.S. Pat. No. 4,168,146 to Grubb et al., U.S. Pat. No.4,366,241 to Tom et al., U.S. Pat. Nos. 6,017,767 and 5,998,220 to Chandler; and U.S. Pat. No. 4,305,924 to Piasio et al.
In competitive or indirect immunoassays, the immobilized component is present in controlled amounts and the mobile component is present in unknown amounts. The unknown amount of mobile component is supplemented with a known amount of the same component that has been tagged by the addition of a measurable constituent which does not interfere with its immunochemical reactive properties. The tag may consist of a radioisotope, a chromophore, a particle, a fluorophor, or an enzyme. The amount of tagged material bound immuno-chemically to the solid phase will depend upon the amount of untagged component in solution competing for the same binding sites. The more of the unknown component present, the less will be the amount of bound tagged component.
Enzyme-based chromatographic assays are used in addition to immunochromatographic assays. These techniques are roughly analogous to immunochromatographic assays, but use an enzymatically-catalyzed reaction instead of an antigen-antibody reaction. The enzymatically-catalyzed reaction frequently generates a detectable product. Other analogous chromatographic assays are known. Although useful, currently available chromatographic techniques using test strips have a number of drawbacks. Many samples, such as fecal samples, contain particulate matter that can color the pores of the chromatographic medium, greatly hindering the detection of the labeling reagents. Other samples, such as blood, contain cells and colored components that make it difficult to read the test. Wet chromatographic medium is also difficult to read because of specular reflection from the chromatography medium.
Sample preparation and waste generation are responsible for other problems with currently available devices and techniques for immunochromatography. The increased prevalence of diseases spread by infected blood and blood fractions, such as AIDS and hepatitis, has exacerbated these problems. The available forms of lateral flow devices have a large portion of their components that are only used for mechanical support of the chromatographic membrane, and are not sealed, therefore making disposal a difficult, expensive and possibly hazardous procedure because of the presumed bio-hazards. Precautions have to be taken so that workers, or people who may inadvertently come into contact with the waste, do not become contaminated.
One common aspect of known devices, particularly in lateral flow technology, is that the assay is read visually, that is, by means of one or more optically readable lines on a test strip held in a carrier, which may have various configurations. There are several limitations or disadvantages to the known optically detected assays. Because they are optical, only surface changes (typically coloration) can be detected. The target analytes may be in the sample solution but of such a low concentration that only relatively few are captured in the capture zone in the porous membrane of the assay. This may provide a faint or even non-optically detectable line, and a false negative reading can result. Typically one end of the test strip is exposed to the sample, normally a fluid of some type, being tested for the particular target analytes of interest. The fluid migrates through the chromatographic medium whereby the analyte with its label is captured and immobilized, while the remaining fluid is absorbed into a medium at the distal end of the assay strip.
Examples of lateral flow assay methods and apparatuses, where the reading is normally conducted optically, are shown in U.S. Pat. Nos. 5,591,645; 5,798,273; 5,622,871; 5,602,040; 5,714,389; 5,879,951; 4,632,901; and 5,958,790.
Still another limitation on chromatographic devices currently available for use by the clinician or technician is their inability to perform quantitative assays. The labeled sandwich at the capture zone, or the decrease of label at the capture zone of a competitive assay, can only be read from the surface of the membrane, so only a relatively small portion of the label is read. Quantitative assessments are really only an estimation based on color intensity of the detection line. Because the prior art assays are optically read, they are subject to contamination by exposure and light-caused degradation. Optical assays also have a limited shelf life.
Another apparatus for detecting target molecules in a liquid phase is shown in U.S. Pat. No. 5,981,297 where magnetizable particles are employed and the output of magnetic field sensors indicates the presence and concentration of target molecules in the sample being tested. Other examples to sense magnetically using physical forces are disclosed in U.S. Pat. Nos. 5,445,970; 5,981,297; and 5,925,573. However, in these devices, the magnet requires relatively high power because the gap where the assay is placed must be wide enough to accommodate the relatively thick assay device.